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Gene expression profiling of long non-coding RNAs between rainbow trout strains on plant-based diets

ABSTRACT: Rainbow trout (Oncorhynchus mykiss) and other salmonids are piscivorous fish. In aquaculture, fish-based feed ingredients are rapidly becoming unsustainable due to increased demand and diminishing supply. Total replacement of fishmeal with plant proteins causes severe intestinal enteritis, leading to reduced growth and lower feed efficiency. Through selective breeding, we have developed a strain of rainbow trout that does not develop distal intestine enteritis when reared on a high soy plant protein-based feed and also shows increased growth compared to other strains. Since central metabolism plays a major role in dietary alterations, liver RNA was examined for differential expression of long non-coding RNAs (lncRNAs) between commercial and selected trout strains when fed an alternative diet. These types of non-coding RNA have been previously shown to have a potential role in regulation of gene expression; however, most identified lncRNAs have not been functionally characterized and/or lack definition in many non-model organisms. Their roles in gene regulation and their responsiveness to dietary changes in trout are to be explored. After three months of rearing on a plant protein-based (PM) diet, liver tissues from a domestic non-selected strain (House Creek; develops enteritis) and the plant-diet tolerant selected strain (ARS-KO; no enteritis) were extracted and prepped for Illumina RNA-seq. Raw reads were screened for quality then assembled into transcripts using Trinity. Assembled transcripts were then submitted to the Annocript pipeline to determine their potential as lncRNAs. RNA-seq reads were mapped to putative lncRNAs. Read-counts were used to assess differential expression between strains.

ORGANISM(S): Oncorhynchus mykiss

PROVIDER: GSE85104 | GEO | 2018/04/10

REPOSITORIES: GEO

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Project description:Through 8 generations of selection, our group has developed a strain of rainbow trout that exhibits high growth rates on an economically and environmentally sustainable all plant protein, high-soy diet. The selected strain also shows superior performance in bacterial and viral disease challenges compared to commercial trout strains, and even a strain specifically selected over many generations for viral and bacterial disease resistance. The selection criteria was strictly focused on performance on plant-based diets, and therefore the physiological mechanisms responsible for the strain’s superior disease resistance remain unresolved. To better characterize the physiological mechanism behind the superior performance of the selected strain we compared the intestinal gene expression of the select strain to that of a commercial control line of trout during an experimental bacterial infection with Flavobacterium psychrophilum (Fp) (CSF 259-93), the causative agent of bacterial cold water disease (BCWD) in salmonids. At 65 days post hatch, all female rainbow trout from the select and commercial strain were stocked separately into four 150L tanks each, at a density of 45 fish per tank. For both strains of trout, three tanks of fish were experimentally infected with Fp by intramuscular injection and one control tank was mock challenged by sham injection. Sampling was conducted at 5 days post challenge (dpc) (Early Infection) and 21 dpc (Late/Recovered Infection). Two intestinal samples from each tank were pooled and two pools from each tank were utilized for RNAseq library preparation. The select strain of trout showed significantly better survival rates (Log-Rank Test, p < 0.0001) over the 21 day infection period, with 70 and 95 % mortality among the select and commercial strain, respectively. Reads from the RNAseq samples were quantified at the transcript level prior to evaluating differential transcript usage and differential gene expression between the strains of trout, infection time points, and disease status.

Project description:Sustainable aquaculture, which entails proportional replacement of fish-based feed sources by plant-based ingredients, is impeded by the poor growth response frequently seen in fish fed high levels of plant ingredients. To evaluate the potential to improve, by means of early nutritional exposure, the growth of fish fed plant-based feed, rainbow trout swim-up fry were fed for 3 weeks either a plant-based diet (diet V, V-fish) or a diet containing fishmeal and fish oil as protein and fat source (diet M, M-fish). After this 3-week nutritional history period, all V- or M-fish received diet M for a 7-month intermediate growth phase. Both groups were then challenged by feeding diet V for 25 days during which voluntary feed intake, growth, and nutrient utilization were monitored (V-challenge). The results of the V-challenge showed a higher growth rate and higher feed intake in fish of nutritional history V compared to M (see PMID:24386155). To identify the molecular mechanisms that govern the enhancement of feed acceptance following early-feed exposure of diet V, we performed microarray expression analysis using swim-up fry collected at the end of early-feeding exposure (diet M and diet V), as well as brain and liver of juveniles sampled at the end of the V-challenge (M-his and V-his), and employing a rainbow trout microarray platform with 8 X 60K probes per slide. Persistently differentially expressed genes were identified by isolating probes that were differentially expressed (fold change ≥ 1.5 and p {nutritional history} ≤ 0.05), both at the end of the early exposure, and in either brain or liver of juvenile trout at the end of the V-challenge. Overall, 1787 (3-week + Brain) and 924 (3-week + Liver) mRNA probes were found to be persistently affected by nutritional history of early-feeding exposure. This positive response is encouraging as a potential strategy to improve the use of plant-based feed in fish, of interest in the field of fish farming and animal nutrition in general. Furthermore, the genes associated with mediating this positive early feeding effect, could serve as potential biomarkers to evaluate increasing acceptability of plant-based diets in fish farming.

Project description:Whirling disease, caused by the pathogen Myxobolus cerebralis, afflicts several salmonid species. Rainbow trout are particularly susceptible and suffer high mortality rates. The disease is persistent and spreading in hatcheries and natural waters of several countries, including the U.S., and the economic losses associated with whirling disease have been extremely large. In this study, gene expression profiling was conducted for resistant and susceptible rainbow trout strains following pathogen exposure with the primary objective of identifying specific genes associated with whirling disease resistance. Several genes were significantly up-regulated in skin following pathogen exposure for both the resistant Hofer and susceptible Trout Lodge rainbow trout strains. For both strains, response to infection appears to be associated with the interferon system. Metallothionein is differentially expressed between the resistant and susceptible strains and is a good candidate for future studies due to its diverse functional roles in inflammation and immune response. The present study has provided the first examination into the genetic basis underlying rainbow trout’s immune response and resistance to the whirling disease pathogen. The identified genes have allowed us to gain initial insight into the molecular mechanisms associated with a salmonid host’s immune response and resistance to whirling disease infection. Keywords: Disease state comparison. 1st comparison: exposed vs control. 2nd comparison: resistant vs susceptible strains Resistant (Hofer) and susceptible (Trout Lodge) rainbow trout strains were exposed to 2,000 triactinomyxons (whirling disease pathogen) per fish. Unexposed controls for each strain were treated identically to exposed other than their lack of pathogen exposure. After twenty-four hours, caudal fin tissue was collected for RNA extraction. Amplified RNA was competitively hybridized (exposed versus control) for each strain. Four biological replicates were used for each experimental condition and a balanced block design was incorporated.

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Gene expression profiling of long non-coding RNAs between rainbow trout strains on plant-based diets

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